PROJECT SUMMARY Lung transplant surgery is the only definitive therapy for end-stage lung disease. While the waiting list of patients eligible for transplant grows, the number of suitable donor organs falls far short of the demand. A significant adverse event that occurs after lung transplantation is primary graft dysfunction (PGD); ischemia- reperfusion (IR) induced cellular injury and tissue degradation play a significant role in PGD. As a community, we throw away ~80% of the potential lung donors out of fear of an adverse PGD outcome. An agent to reduce ischemic damage to organs and repair them prior to transplantation will represent a technology that can preserve lung integrity, resuscitate marginal donor organs, and improve the overall positive impact of transplantation. Previously we identified MG53 that functions as an essential component of cell membrane repair machinery. Genetic ablation of MG53 (mg53-/- mice) leads to increased susceptibility of the lung to stress induced injuries. Transgenic mice with sustained elevation of MG53 in the bloodstream (tPA-MG53 mice) live a healthy lifespan with enhanced regenerative capacity following injury to multiple organs. The recombinant human MG53 protein (rhMG53) has great potential in prevention and protection of injuries to the lung, heart and kidney in rodent and large animal models. Using ex-vivo lung perfusion (EVLP) as a platform for delivering therapeutic proteins, we show that rhMG53 improves lung donor integrity by mitigating injuries to both alveolar and endothelia cells. This project builds on the scientific premise that targeting the elemental process of cell membrane repair represents a potential novel means to preserve the quality of the lung donor, which shall improve the outcome of lung transplantation (e.g. reduction of PGD). We will conduct lung transplants between donor and recipient mg53-/- and tPA-MG53 mice to define the physiologic role of MG53 in lung protection associated with IR injury (Aim 1). We will use cellular, molecular and live cell imaging tools to derive a mechanistic base for how circulating MG53 preserves the endothelial cell integrity, improves cell survival and reduces inflammation during IR; these studies will also establish a set of bio-markers modulated by rhMG53 that can quantifiably predict lung injury prior to transplantation (Aim 2). In Aim 3, we will establish the quality controls for production and formulation of rhMG53 for pulmonary applications, and use EVLP as a delivery vehicle to test the efficacy of rhMG53 in rescuing the injured lung allograft derived from pigs and humans. Finally, we will test in a porcine model of lung transplantation whether addition of rhMG53 during the preservation process or after transplantation can have beneficial effects on prevention of PGD.